Orthotopic transplantation of immortalized mesencephalic progenitors (CSM14.1 cells) into the substantia nigra of hemiparkinsonian rats induces neuronal differentiation and motoric improvement

Abstract

Neural progenitor cell grafting is a promising therapeutic option in the treatment of Parkinson's disease. In previous experiments we grafted temperature-sensitive immortalized CSM14.1 cells, derived from the ventral mesencephalon of E14-rats, bilaterally in the caudate putamen of adult hemiparkinsonian rats. In these studies we were not able to demonstrate either a therapeutic improvement or neuronal differentiation of transplanted cells. Here we examined whether CSM14.1 cells grafted bilaterally orthotopically in the substantia nigra of hemiparkinsonian rats have the potential to differentiate into dopaminergic neurons. Adult male rats received 6-hydroxydopamine into the right medial forebrain bundle, and successful lesions were evaluated with apomorphine-induced rotations 12 days after surgery. Two weeks after a successful lesion the animals received bilateral intranigral grafts consisting of either about 50 000 PKH26-labelled undifferentiated CSM14.1 cells ( n = 16) or a sham-graft ( n = 9). Rotations were evaluated 3, 6, 9 and 12 weeks post-grafting. Animals were finally perfused with 4% paraformaldehyde. Cryoprotected brain slices were prepared for immunohistochemistry using the freeze-thaw technique to preserve PKH26-labelling. Slices were immunostained against neuronal epitopes (NeuN, tyrosine hydroxylase) or glial fibrillary acidic protein. The CSM14.1-cell grafts significantly reduced the apomorphine-induced rotations 12 weeks post-grafting compared to the sham-grafts ( P < 0.05). There was an extensive mediolateral migration (400-700 µ m) of the PKH26-labelled cells within the host substantia nigra. Colocalization with NeuN or glial fibrillary acidic protein in transplanted cells was confirmed with confocal microscopy. No tyrosine hydroxylase-immunoreactive grafted cells were detectable. The therapeutic effect of the CSM14.1 cells could be explained either by their glial cell-derived neurotrophic factor-expression or their neural differentiation with positive effects on the basal ganglia neuronal networks.

The Proteome of the Differentiating Mesencephalic Progenitor Cell Line CSM14.1In Vitro

“…In vivo NeuN-positive CSM14.1 cells were detectable after transplantation into the striatum of neonatal rats [10] or into the substantia nigra of adult hemiparkinsonian rats [21]. Assuming that CSM14.1 cells at permissive culture conditions (33°C, 10% FCS) are undifferentiated, these cells should not show immunoreactivity against NeuN, which could be demonstrated in the present study.…”

Differential Proteomics of the Cerebral Cortex of Juvenile, Adult and Aged Rats: An Ontogenetic Study

“…Several studies [24][25][26] focus on the transplantation of immortalized progenitor cells in 6-OHDA Hemiparkinson animal models [24,25] and the differences in the abundance of this area of the brain at different points of development [26]. Hovakimyan et al [24] and Haas et al [25] as well as Schwarz and Freed [27] showed that the micro-environment of neonatal brains seems to be important for the differentiation of transplanted neuronal progenitors. Therefore, a further characterization of the abundance patterns of proteins during development in the Cx of the rat brain might provide a better understanding of neuronal development.…”

Immortalization of neuronal progenitors using SV40 large T antigen and differentiation towards dopaminergic neurons

“…Further differentiation of these cells led to the time-dependent expression of TH and aldehyde dehydrogenase 2 (AHD2) at translational level [45]. Transplantation of the cells in hemiparkinsonian animals resulted in the reduction of apomorphine-induced rotation [46,47] and no tumour formation was observed in transplanted grafts or the surrounding host tissue [48]. …”

“…Likewise, conditionally immortalized cells like CSM14.1 have also been shown to reduce apomorphine-induced rotation in hemiparkinsonian animals [48]. Palmer et al .…”

“…Likewise, immortalized cell lines of olfactory ensheathing glia with SV40 LT upon transplantation in animal model of spinal cord injury shows that there was no LT expression in the grafts after 4 weeks of transplantation and the animals’ recovered sensory and motor function [118]. CSM14.1 cell line on transplantation in hemiparkinsonian rats did not form tumours and were not able to express SV40 LT [48]. In conditionally immortalized chromaffin cell line with SV40 LT derived from E17 rat adrenal and neonatal bovine, adrenal cells were not exhibiting LT expression when transplanted in the lumbar subarachnoid space of spinal cord [106,108].…”